Title

Authors

Date of Completion

January 2004

Keywords

Biology, Neuroscience

Degree

Ph.D.

Abstract

Fast ionotropic inhibitory neurotransmission, mediated by presynaptic GABAergic terminals and correctly positioned postsynaptic GABAA receptors (GABAARs), occurs in about 30–40% of all synapses in the CNS. However, little is known about the specific control mechanisms responsible for patterned expression, assembly, trafficking, and targeting of GABAA receptors to GABAergic synapses or extrasynaptic domains within a single neuron. In this study, a cultured hippocampal neuron (CHN) model was used to examine the distribution and patterning of 13 different GABAAR subunit isoforms from 4 distinct subunit classes normally expressed in the brain. ^ The group of experiments in this thesis shows that CHNs, as in the intact hippocampus, express and postsynaptically concentrate (cluster) GABA AR subtypes containing the α1–3, α 5, β1–3, γ2S, γ2L and γ 3 subunit isoforms. This postsynaptic clustering of GABAARs in CHNs primarily occurs apposed to inhibitory presynaptic contact and contains receptors with the γ2 or γ3 subunit. However, when presynaptic GABAergic terminals are absent from the local dendritic environment, significant GABAAR clustering occurs apposed to excitatory synaptic contact forming ‘mismatched’ synapses. GABAAR clustering is particularly pronounced within the synapses onto the axon initial segment (AIS) of pyramidal cells in the brain, a phenomenon that also occurs in the AIS of CHNs independently of GABAergic or glutamatergic terminal phenotype. However, the coexistence of two different terminal phenotypes does not seem possible within this subcellular structure, due to the hierarchy of organizational signals created by correctly matched GABAergic synapses vs. mismatched synapses. Thus, we show that competition between GABAergic and glutamatergic terminals ultimately determines the postsynaptic localization of the GABAAR subtypes with a capacity for clustering. Lastly, this work addressed a molecular mechanism of GABAAR clustering. CHNs, as in the brain, are capable of expressing non-clustered GABAAR subtypes containing the δ subunit. We have examined expression of chimeric subunits in which the large intracellular loops (IL) of γ2S and δ subunits were exchanged. We show that while the γ2 IL is both necessary and central to the clustering process, it is not sufficient for clustering when incorporated into the δ subunit. Thus, there are additional requirements other than the γ2-IL for receptor clustering that are as yet unidentified. ^